Bone marrow and peripheral blood progenitor cell transplantation are clinical procedures in which donor bone marrow or peripheral blood cells are transplanted into a recipient for the reconstitution of the recipient's lymphohematopoietic system. Prior to the transplant, the recipient's own blood system is either naturally deficient or intentionally destroyed by agents such as irradiation. In cases where the recipient is a cancer patient, ablative therapy is often used as a form of cancer treatment which also destroys the cells of the lymphohematopoietic system. The success rate of this procedure depends on a number of critical factors, which include the number of hematopoietic progenitor cells in the donor cell preparation, matching between donor and recipient at the major histocompatibility complex (MHC) which encodes products that induce graft rejection, and conditioning of the recipient prior to transplantation.
Tissue typing technology has ushered in dramatic advances in the use of allogeneic bone marrow cells as a form of therapy in patients with deficient or abnormal hematopoiesis. Conditioning of a recipient can be achieved by total body or total lymphoid irradiation. However, methods to enrich for the hematopoietic progenitor cell in a donor cell preparation are still not fully perfected. A pluripotent progenitor cell is believed to be capable of self-renewal and differentiation into blood cells of various lineages including lymphocytes, granulocytes, macrophages/monocytes, erythrocytes and megakaryocytes (Ikuta et al., 1992, Ann. Rev. Immunol. 10:759). Recent studies have shown that progenitor cells reside in the CD34.sup.+ cell population in that anti-CD34 antibody-purified CD34.sup.+ cells can repopulate all hematopoietic cell types in lethally-irradiated patients. The mechanism by which a progenitor cell commits to a specific cell lineage has not been fully elucidated. However, it is clear that such events must, in part, be influenced by a variety of growth and differentiation factors that specifically regulate hematopoiesis. Other factors which are not yet identified may also be involved (Metcalf, 1989, Nature 339:27). The commonly known hematopoietic factors include erythropoietin (EPO), granulocyte/macrophage colony stimulating factor (G/M-CSF), granulocyte colony-stimulating factor (G-CSF), macrophage colony-stimulating (M-CSF), interleukin 1-12 (IL-1 to IL-12), and progenitor cell factor (SCF).
The ability to enrich for CD34.sup.+ cells is critical to the application of bone marrow transplantation as a form of therapy for hematopoietic disorders. Neoplastic transformation, immunodeficiency, genetic abnormalities, and even viral infections can all affect blood cells of different lineages and at different stages of development. Bone marrow transplantation provides a potential means for treating all such disorders. In addition, although bone marrow transplantation may not be used as a direct form of treatment for solid tumors, it provides an important means of maintaining survival of patients following various ablative therapeutic regimens. Current conventional therapy utilizes whole bone marrow harvested from the iliac crest but this approach has certain limitations. For example, bone marrow progenitor cells are present in extremely low numbers, and bone marrow aspiration involves painful invasive procedures.
If the bone marrow cells or other progenitor cell source contain contaminating tumor cells that must be purged prior to re-infusion in an autologous setting, the large number of total cells with a low percentage of CD34.sup.+ cells makes it technically difficult to perform adequate purging of tumor cells. Thus, there remains a need for a simple method for enriching CD34.sup.+ progenitor cells from a cell mixture containing higher numbers of these cells that are amenable to efficient purging of residual tumor cells for use in subsequent transplantation.
In an effort to address these problems, investigators have focused on the use of anti-CD34 antibodies. Such procedures involve positive selection, such as the passage of white blood cells over a column containing anti-CD34 antibodies or binding of cells to magnetic bead-conjugated anti-CD34 antibodies or by panning on anti-CD34-coated plates, and collecting the bound cells. However, the affinity based methods have practical limitations in that they are not reusable and are costly.
Alternative methods for enriching hematopoietic progenitor cells have been reported which utilized various forms of density gradient centrifugation (Olofsson et al., 1980, Scan. J. Haematol. 24:254; Ellis et al., 1984, J. Immunol. Meth. 66:9; Lasky and Zanjani, 1985, Exp. Hematol. 13:680; Martin et al., 1986, Brit. J. Haematol. 63:187). However, all reported methods use agar colony assays to identify hematopoietic progenitor cells after enrichment. It is known that the progenitor assays only detect committed precursor cells which occupy less than 1% of the CD34.sup.+ population. It is therefore uncertain whether these methods can in fact enrich for the early progenitor cells or stem cells which can permanently engraft and reconstitute a lymphohematopoietic system, as they have not been tested clinically. Furthermore, there is no indication from the published reports that any of these procedures are able to obtain adequate numbers of cells for clinical use.